Calcitonin Gene-Related Peptide Attenuates Hyperoxia-Induced Oxidative Damage in Alveolar Epithelial Type II Cells Through Regulating Viability and Transdifferentiation.

As a stem cell of alveolar epithelium, the physiological status of alveolar epithelium type II cells (AECII) after hyperoxia exposure is closely related to the occurrence of hyperoxia-induced lung injury and the restoration of normal morphological function of damaged alveolar epithelium. However, the relevant mechanisms involved are not very clear. Therefore, this study aimed to explore the effect of calcitonin gene-related peptide (CGRP) on AECII exposed to hyperoxia and its potential mechanisms. The AECII viability was detected using MTT assay. The malondialdehyde (MDA) level and superoxide dismutase (SOD) activity were detected by spectrophotometry. The transdifferentiation capacity of AECII was evaluated by flow cytometry. The expression levels of Notch1, Hes, HERP, and AECII markers were detected using immunohistochemistry and/or RT-qPCR or immunofluorescence. ELISA was used for the determination of inflammatory markers. The results showed that CGRP significantly promoted cell viability, and markedly suppressed hyperoxia-induced transdifferentiation of AECII; these biological alterations were coincided with decreased MDA level, increased SOD activity, and activated Notch signaling pathway (upregulated expression levels of Notch1, Hes, and HERP). Notably, the in vitro effects of CGRP on Notch signaling pathway were further investigated in animal model, and the HE staining results showed that CGRP reduced in vivo oxidative injury and inflammation in hyperoxia-treated AECII through the promotion of structural and functional regeneration, accompanied by elevated Notch1 expression and activated Notch signaling cascade as shown by immunohistochemistry and QPCR, respectively. Immunohistochemistry of APQ-5 and SPC indicated that CGRP reversed the transdifferentiation of AECIIs in vivo. Our current results were consistent across both in vitro and in vivo settings, and provide a new direction for the prevention and treatment of bronchopulmonary dysplasia (BPD).

Knockdown of Long Noncoding RNAs of Maternally Expressed 3 Alleviates Hyperoxia-Induced Lung Injury via Inhibiting Thioredoxin-Interacting Protein-Mediated Pyroptosis by Binding to miR-18a.

Long-term hyperoxia exposure may cause lung damage with characteristic inflammation. Long noncoding RNA of maternally expressed 3 (MEG3) is up-regulated in lung tissues exposed to hyperoxia; however, the underlying mechanism is unclear. Hyperoxia-induced cells and mouse models were used to study these mechanisms. Molecular assays were used to detect cell viability, cytotoxicity, and expression of miR-18a, MEG3, and inflammatory cytokines. The interaction among MEG3, miR-18a, and thioredoxin-interacting protein (TXNIP) was verified; and pyroptosis-related proteins were analyzed. The in vivo model was established by exposing MEG3 knockdown mice to hyperoxia. Hematoxylin and eosin staining was used to assess pathologic alterations of lung tissues. Hyperoxia suppressed cell viability, induced cell damage, and exacerbated the secretion of IL-1ß and IL-18. Hyperoxia inhibited miR-18a, with increased expression of MEG3, TXNIP, and nonobese diabetic-like receptor family pyrin domain containing 3 (NLRP3). MEG3 aggravated TXNIP expression by binding to miR-18a. Knockdown of MEG3 rescued hyperoxia-induced pyroptosis by up-regulating miR-18a. Furthermore, knockdown of MEG3 inhibited NLRP3 inflammasome activity and caspase-1 signaling by miR-18a. In vivo knockdown of MEG3 and overexpression of miR-18a relieved hyperoxia-induced lung injury via restraining NLRP3 inflammasome-mediated pyroptosis, whereas miR-18a inhibition reversed these effects. In conclusion, knockdown of MEG3 inhibits pyroptosis to alleviate hyperoxia lung injury by suppressing NLRP3 inflammasome and caspase-1 signaling via regulating miR-18a-TXNIP axis.